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Long-distance transport cadmium (Cd) from roots to shoots is a key factor for Cd phytoremediation. Our previous study indicated that heavy metal P1B2-ATPases, IlHMA2, was involved in improving the accumulation of Cd via mediated long-distance transport Cd, contributing to the phytoremediation in Cd accumulator Iris lactea. However, whether the overexpression of IlHMA2 could enhance the accumulation and tolerance to Cd remains unclear in plants. Here, we generated transgenic tobacco overexpressing IlHMA2 and tested its effect on the translocation and accumulation of Cd and zinc (Zn), as well as the physio-biochemical characteristics under 50 mg/L Cd exposure. The overexpression of IlHMA2 significantly increased Cd concentrations in xylem saps, resulting in enhanced root-to-shoot Cd translocation compared with wild-type. Meanwhile, overexpressing IlHMA2 promoted Zn accumulations, accompanied by elevating proline contents and antioxidant enzyme activity (SOD, POD, and CAT) to diminish the overproduction of ROS in transgenic tobacco. These pieces of evidence suggested that higher Zn concentrations and lower ROS levels could tremendously alleviate Cd toxicity for transgenic tobacco, thereby improving the growth and tolerance. Overall, the overexpression of IlHMA2 could facilitate Cd accumulation and enhance its tolerance in tobacco exposed to Cd contaminations. This would provide a valuable reference for improving Cd phytoremediation efficiency. [ABSTRACT FROM AUTHOR]

Miscanthus spp. show excellent application prospects due to its bioenergy potential and multiple ecological services. Annual N export with biomass harvest from Miscanthus, even without fertilizer supplement, do not reduce soil N levels. The question arises regarding how Miscanthus can maintain stable soil N levels. Metagenomic strategies were used to reveal soil N‐cycling‐related microbiome and their functional contributions to processes of soil N‐cycling based on the comparison among the bare land, cropland, 10‐year Miscanthus × giganteus, and 15‐year Miscanthus sacchariflorus fields. The results showed that, after long‐term cropland‐to‐Miscanthus conversion (LCMC), 16 of 21 bacterial phyla and all the archaeal phyla exhibited significant changes. Soil microbial denitrification and nitrification functions were significantly weakened, and N fixation (NF) was significantly enhanced. The biosynthesis of amino acids, especially alanine, aspartate, and glutamate metabolism, in soil N‐cycling‐related microbiome was dramatically promoted. The genus Anaeromyxobacter contributed largely to the NF process after LCMC. Variations in the soil available potassium, available N, organic C, and total N contents drove a functional shift of soil microbiome from cropland to Miscanthus pattern. We conclude that Miscanthus can recruit Anaeromyxobacter communities to enhance NF benefiting its biomass sustainability and soil N balance. [ABSTRACT FROM AUTHOR]

The extensive applications of graphene oxide (GO) inevitably lead to entry into the natural aquatic environment. However, information on its toxicity to emergent plants is still lacking. In this study, an emergent plant, Iris pseudacorus, was exposed to GO (1, 20, 80, and 140 mg·L−1) under hydroponic conditions for 15 weeks. Changes in plant growth were assessed by analyzing plant biomass and photosynthetic pigment contents; the photosynthesis response was verified by measuring chlorophyll a fluorescence; and the nutrient levels of the plant were evaluated. Results showed that GO at 20–140 mg·L−1 significantly increased plant dry weight by 37–84% and photosynthetic pigment contents by 26–178%, and 80 mg·L−1 was the optimal concentration. PSII activity, adjustment capacities of electron transport in PSII, the grouping or energetic connectivity between PSII units, light energy conversion efficiency, photosynthesis performance indexes (by 11–51%), and contents of several nutrient elements (N, Fe, and Cu) were increased by 49–69%, 34–84%, and 11–38%, respectively. These findings indicate that GO can enhance plant growth by promoting plant photosynthesis performance and improving plant nutrient levels, and has great application potential in promoting the growth and development of this emergent plant as a phytoremediation agent. [ABSTRACT FROM AUTHOR]

The bioenergy crop switchgrass (Panicum virgatum L.) has been recognized as friendly to the soil of cultivated land depending on the previous land use types and management practices. However, the effects of switchgrass establishment on soil properties at a broader depth when it is harvested annually without any fertilization in northern China largely remain unknown. To explore the impacts of unfertilized switchgrass on soil physical and chemical properties, 0–100 cm soil samples were collected from 7-year cropland-to-switchgrass conversion and the bare land (control). The results showed that switchgrass establishment increased soil total and capillary porosity, CFU numbers of the microbial communities (fungi, bacteria, and actinomycetes), contents of microbial biomass (carbon, nitrogen, and phosphorus), and water-soluble organic carbon, and decreased soil bulk density, mostly at 0–60 cm depths, compared to the control values. Notably, the annual harvest of switchgrass insignificantly increased soil total and available nitrogen contents and slightly reduced available phosphorus and potassium contents. In conclusion, long-term cropland conversion to unfertilized switchgrass could ameliorate soil properties and does not cause soil depletion. The output of this study could inspire governments and farmers to make large-scale use of switchgrass in the ecological restoration of abandoned cropland in north China. [ABSTRACT FROM AUTHOR]

The recretohalophyte Karelinia caspia is of forage and medical value and can remediate saline soils. We here assess the contribution of primary/secondary metabolism to osmotic adjustment and ROS homeostasis in Karelinia caspia under salt stress using multi‐omic approaches. Computerized phenomic assessments, tests for cellular osmotic changes and lipid peroxidation indicated that salt treatment had no detectable physical effect on K. caspia. Metabolomic analysis indicated that amino acids, saccharides, organic acids, polyamine, phenolic acids, and vitamins accumulated significantly with salt treatment. Transcriptomic assessment identified differentially expressed genes closely linked to the changes in above primary/secondary metabolites under salt stress. In particular, shifts in carbohydrate metabolism (TCA cycle, starch and sucrose metabolism, glycolysis) as well as arginine and proline metabolism were observed to maintain a low osmotic potential. Chlorogenic acid/vitamin E biosynthesis was also enhanced, which would aid in ROS scavenging in the response of K. caspia to salt. Overall, our findings define key changes in primary/secondary metabolism that are coordinated to modulate the osmotic balance and ROS homeostasis to contribute to the salt tolerance of K. caspia. [ABSTRACT FROM AUTHOR]

Miscanthus spp. are increasingly cultivated in cropland worldwide due to their bioenergy potential and multiple ecological services. Effects of long‐term cropland‐to‐Miscanthus conversion without N fertilizer on soil microbiome and N cycling largely remain unknown. We aimed to explore the effects of Miscanthus conversion on soil microbiome and N cycling over a 15‐year period. We analyzed diversity, composition, and abundance of bacterial and archaeal communities using 16S rRNA amplicon sequencing, and abundances of N‐cycling‐related genes using quantitative polymerase chain reaction of 0–10 cm soils collected from bare land, cropland, 10‐year Miscanthus × giganteus, and 15‐year Miscanthussacchriflorus land in Beijing. Conversion decreased soil sand and micro‐aggregate proportion, nitrate N (NiN), available phosphorus levels, conductivity, temperature, and pH, while increasing proportion of soil clay and macro‐aggregate (MAA), soil organic C (SOC), available N (AN), exchangeable Mg2+ (EMg2+), and available potassium (AK) contents as well as microbial C/N. Consequently, diversity, composition, and abundance of soil bacterial community exhibited larger changes than those values of archaeal community after conversion. Soil AP, EMg2+, AK, and SOC were key factors in shifting microbiome from the cropland to Miscanthus pattern. Moreover, abundances of bacterial and archaeal communities and the N fixer gene nifH increased, whereas that of the bacterial ammonia monooxygenase gene decreased. The copies of other N‐cycling‐related genes in the two Miscanthus lands seemed similar to those values of cropland. The nifH copies negatively correlated with soil NiN and positively correlated with AN, EMg2+, ECa2+, SOC, AK, and MAA. We conclude that changes in soil microbiome pattern induced by the variation of soil properties enhance microbial N fixation potential, maintaining stable N levels and robust N cycling with lower N leakage risk after conversion. These results should inspire farmers and governments to large‐scale use Miscanthus on marginal cropland in Northern China. [ABSTRACT FROM AUTHOR]

Abstract Background Lignocellulose is the most abundant biomass on earth. However, biomass recalcitrance has become a major factor affecting biofuel production. Although cellulose crystallinity significantly influences biomass saccharification, little is known about the impact of three major wall polymers on cellulose crystallization. In this study, we selected six typical pairs of Miscanthus samples that presented different cell wall compositions, and then compared their cellulose crystallinity and biomass digestibility after various chemical pretreatments. Results A Miscanthus sample with a high hemicelluloses level was determined to have a relatively low cellulose crystallinity index (CrI) and enhanced biomass digestibility at similar rates after pretreatments of NaOH and H2SO4 with three concentrations. By contrast, a Miscanthus sample with a high cellulose or lignin level showed increased CrI and low biomass saccharification, particularly after H2SO4 pretreatment. Correlation analysis revealed that the cellulose CrI negatively affected biomass digestion. Increased hemicelluloses level by 25% or decreased cellulose and lignin contents by 31% and 37% were also found to result in increased hexose yields by 1.3-times to 2.2-times released from enzymatic hydrolysis after NaOH or H2SO4 pretreatments. The findings indicated that hemicelluloses were the dominant and positive factor, whereas cellulose and lignin had synergistic and negative effects on biomass digestibility. Conclusions Using six pairs of Miscanthus samples with different cell wall compositions, hemicelluloses were revealed to be the dominant factor that positively determined biomass digestibility after pretreatments with NaOH or H2SO4 by negatively affecting cellulose crystallinity. The results suggested potential approaches to the genetic modifications of bioenergy crops.

Aim: The aim of this study was to understand the effects of revegetation on the diversity of bacteria and fungi in soil by sowing a single species and exploring the underlying mechanism. Location: Beijing, China. Taxon: Plants and Microbes. Methods: In a short‐term ecological restoration experiment, one natural recovery treatment and three seed sowing treatments were chosen to assess their effects on the alteration of fungal and bacterial diversity. Plant species richness, abundance, and height were investigated. The diversity of fungi and bacteria was analyzed by high‐throughput sequencing technologies. Linear mixed‐effects model analysis was used to examine the effects of different restoration methods on biodiversity and ecosystem functions. Pearson's correlation analysis, analysis of covariance, and structural equation modeling (SEM) were used to examine the relationship between biodiversity and environmental factors. Results: Species richness and the Shannon–Wiener Index (H′) of plants in the sown treatments were lower than in the natural recovery treatment, especially with sowing of Medicago sativa L. Similarly, the sum of the observed species and H′ of fungi and bacteria significantly decreased in the sown treatments. Moreover, plant density, community coverage, and soil moisture increased markedly, while soil bulk density decreased in the sown treatments. Importantly, SEM showed that sown treatments reduced the diversity of plants through increasing plant density, while it decreased the diversity of fungi and bacteria through decreasing the plant diversity and increasing soil moisture. Main conclusions: Our findings confirm that ecological restoration by sowing could improve soil conditions, but may be unfavorable to the amelioration of soil microbial diversity in the short‐term. Restoration practitioners should consider long‐term studies on the dynamics of biodiversity in the above‐ and belowground after revegetation by native species to achieve goals related to biodiversity conservation. [ABSTRACT FROM AUTHOR]

Switchgrass is a photoperiod-sensitive energy grass suitable for growing in the marginal lands of China. We explored the effects of extended photoperiods of low-irradiance light (7 μmol·m-2·s-1, no effective photosynthesis) on the growth, the biomass dry weight, the biomass allocation, and, especially, the stem digestibility and cell wall characteristics of switchgrass. Two extended photoperiods (i.e., 18 and 24 h) were applied over Alamo. Extended light exposure (18 and 24 h) resulted in delayed heading and higher dry weights of vegetative organs (by 32.87 and 35.94%, respectively) at the expense of reducing the amount of sexual organs (by 40.05 and 50.87%, respectively). Compared to the control group (i.e., natural photoperiod), the yield of hexoses (% dry matter) in the stems after a direct enzymatic hydrolysis (DEH) treatment significantly increased (by 44.02 and 46.10%) for those groups irradiated during 18 and 24 h, respectively. Moreover, the yield of hexoses obtained via enzymatic hydrolysis increased after both basic (1% NaOH) and acid (1% H2SO4) pretreatments for the groups irradiated during 18 and 24 h. Additionally, low-irradiance light extension (LILE) significantly increased the content of non-structural carbohydrates (NSCs) while notably reducing the lignin content and the syringyl to guaiacyl (S/G) ratio. These structural changes were in part responsible for the observed improved stem digestibility. Remarkably, LILE significantly decreased the cellulose crystallinity index (CrI) of switchgrass by significantly increasing both the arabinose substitution degree in xylan and the content of ammonium oxalate-extractable uronic acids, both favoring cellulose digestibility. Despite this LILE technology is not applied to the cultivation of switchgrass on a large scale yet, we believe that the present work is important in that it reveals important relationships between extended day length irradiations and biomass production and quality. Additionally, this study paves the way for improving biomass production and digestibility via genetic modification of day length sensitive transcription factors or key structural genes in switchgrass leaves. [ABSTRACT FROM AUTHOR]

Tachysurus vachellii are commercially important edible fish due to delicious taste, little bone in muscle, and high nutritional value especially in Asia. The complete mitochondrial genome of Tachysurus vachellii has been sequenced. The mitochondrial genome is 16,529 bp in length, with the base composition of 31.6% A, 26.6% T, 26.9% C, and 14.9% G, containing two ribosomal RNA genes, 13 protein-coding genes, 22 transfer RNA genes, and a major non-coding control region (D-loop region). The gene order and orientation are similar with some typical fish species. The data will provide useful molecular information for phylogenetic studies concerning T. vachellii and its related species. [ABSTRACT FROM AUTHOR]

In this study, tassels of Cave-in-Rock (upland) and Alamo (lowland) were removed at or near tassel emergence to explore its effects on biomass production and quality. Tassel-removed (TR) Cave-in-Rock and Alamo both exhibited a significant (P<0.05) increase in plant heights (not including tassel length), tiller number, and aboveground biomass dry weight (10% and 12%, 30% and 13%, 13% and 18%, respectively by variety) compared to a control (CK) treatment. Notably, total sugar yields of TR Cave-in-Rock and Alamo stems increased significantly (P<0.05 or 0.01) by 19% and 19%, 21% and 14%, 52% and 18%, respectively by variety, compared to those of control switchgrass under 3 treatments by direct enzymatic hydrolysis (DEH), enzymatic hydrolysis after 1% NaOH pretreatment (EHAL) and enzymatic hydrolysis after 1% H2SO4 pretreatment (EHAC). These differences were mainly due to significantly (P<0.05 or 0.01) higher cellulose content, lower cellulose crystallinity indexes (CrI) caused by higher arabinose (Ara) substitution in xylans, and lower S/G ratio in lignin. However, the increases of nitrogen (N) and sulphur (S) concentration negatively affects the combustion quality of switchgrass aboveground biomass. This work provides information for increasing biomass production and quality in switchgrass and also facilitates the inhibition of gene dispersal of switchgrass in China. [ABSTRACT FROM AUTHOR]

Plant lignin is one of the major wall components that greatly contribute to biomass recalcitrance for biofuel production. In this study, total 79 representative Miscanthus germplasms were determined with wide biomass digestibility and diverse monolignol composition. Integrative analyses indicated that three major monolignols (S, G, H) and S/G ratio could account for lignin negative influence on biomass digestibility upon NaOH and H2SO4 pretreatments. Notably, the biomass enzymatic digestions were predominately affected by the non-KOH-extractable lignin and interlinked-phenolics, other than the KOH-extractable ones that cover 80% of total lignin. Furthermore, a positive correlation was found between the monolignols and phenolics at p<0.05 level in the non-KOH-extractable only, suggesting their tight association to form the minor wall-networks against cellulases accessibility. The results indicated that the non-KOH-extractable lignin-complex should be the target either for cost-effective biomass pretreatments or for relatively simply genetic modification of plant cell walls in Miscanthus. [ABSTRACT FROM AUTHOR]

Plant lignin is one of the major wall components that greatly contribute to biomass recalcitrance for biofuel production. In this study, total 79 representative Miscanthus germplasms were determined with wide biomass digestibility and diverse monolignol composition. Integrative analyses indicated that three major monolignols (S, G, H) and S/G ratio could account for lignin negative influence on biomass digestibility upon NaOH and H2SO4 pretreatments. Notably, the biomass enzymatic digestions were predominately affected by the non-KOH-extractable lignin and interlinked-phenolics, other than the KOH-extractable ones that cover 80% of total lignin. Furthermore, a positive correlation was found between the monolignols and phenolics at p<0.05 level in the non-KOH-extractable only, suggesting their tight association to form the minor wall-networks against cellulases accessibility. The results indicated that the non-KOH-extractable lignin-complex should be the target either for cost-effective biomass pretreatments or for relatively simply genetic modification of plant cell walls in Miscanthus. [ABSTRACT FROM AUTHOR]

• Vegetated ditch can effectively intercept nitrogen in farmland runoff. • The 15N stable isotope tracer method was used to analyze the fate of vegetated ditch intercepting farmland runoff nitrogen. • The nitrogen remaining in the L. perenne and E. repens ditch beds was almost completely absorbed by the plants after 60 d of growth. During periods of heavy precipitation, farmland drainage ditches are the main routes for farmland nitrogen flowing into surface waters in northern China. Vegetated ditches can be engineered to mitigate N loss from agricultural runoff. We used a simulation study to evaluate the interception of nitrogen in a ditch without vegetation and three vegetated ditches. The 15N stable isotope tracer method was used to analyze the fate of ditch intercepting farmland runoff nitrogen. The three vegetated ditches were planted with Lolium perenne L., Bromus inermis Leyss. and Elytrigia repens (L.) Nevski. The results showed that the interception rates of nitrogen in the three vegetated ditches were higher than in the ditch without vegetation. The L. perenne ditch had the lowest interception rates of nitrogen among the three vegetated ditches. The interception of nitrogen by the three vegetated ditches was primarily achieved by the capture of nitrogen runoff in the soil ditch bed, which allowed water infiltration. Nitrogen retained in the ditch bed soil of the ditch without vegetation was enriched in the 0–10 cm soil layer from surface mainly. However, in the three vegetated ditches, it was transported to the 50 cm soil layer. All of the nitrogen retained in the vegetated ditches was absorbed by the L. Perenne and E. repens , and the interception of nitrogen was affected by plant species and growth stage in the vegetated ditch. [ABSTRACT FROM AUTHOR]

Polyploidization is regularly regarded as an important tool for the generation of innovative germplasm resources and plant breeding. In this study, autopolyploid switchgrass lines were produced from calli derived from immature inflorescence of tetraploid switchgrass ( Panicum virgatum L.), both lowland cv. Alamo and Kanlow, and upland cv. Ranlow, through colchicine treatments. Ploidy levels of plantlets regenerated from the calli were screened by flow cytometry. Growth characteristics of the tetraploid and octoploid switchgrass cultivars were further investigated in a field experiment. Compared with the tetraploid plants of the three switchgrass cultivars, the corresponding octoploid plants showed significant reduction in plant height, number of tillers per plant and fresh matter, but an increase in leaf length and width. [ABSTRACT FROM AUTHOR]

Pennisetum alopecuroides is an important forage grass resource, which plays a vital role in ecological environment improvement. Therefore, the acquisition of P. alopecuroides genome resources is conducive to the study of the adaptability of Pennisetum species in ecological remediation and forage breeding development. Here we assembled a P. alopecuroides cv. 'Liqiu' genome at the chromosome level with a size of approximately 845.71 Mb, contig N50 of 84.83Mb, and genome integrity of 99.13% as assessed by CEGMA. A total of 833.41-Mb sequences were mounted on nine chromosomes by Hi-C technology. In total, 60.66% of the repetitive sequences and 34,312 genes were predicted. The genomic evolution analysis showed that P. alopecuroides cv. 'Liqiu' was isolated from Setaria 7.53-13.80 million years ago and from Cenchrus 5.33-8.99 million years ago, respectively. The whole-genome event analysis showed that P. alopecuroides cv. 'Liqiu' underwent two whole-genome duplication (WGD) events in the evolution process, and the duplication events occurred at a similar time to that of Oryza sativa and Setaria viridis . The completion of the genome sequencing of P. alopecuroides cv. 'Liqiu' provides data support for mining high-quality genetic resources of P. alopecuroides and provides a theoretical basis for the origin and evolutionary characteristics of Pennisetum ., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Teng, Guo, Liu, Guo, Xu, Hou, Teng, Zhang, Zhao, Yue, Wen, Wu and Fan.)

The halophyte Karelinia caspia has not only fodder and medical value but also can remediate saline-alkali soils. Our previous study showed that salt-secreting by salt glands is one of main adaptive strategies of K. caspia under high salinity. However, ROS scavenging, ion homeostasis, and photosynthetic characteristics responses to high salinity remain unclear in K. caspia . Here, physio-biochemical responses and gene expression associated with ROS scavenging and ions transport were tested in K. caspia subjected to 100-400 mM NaCl for 7 days. Results showed that both antioxidant enzymes (SOD, APX) activities and non-enzymatic antioxidants (chlorogenic acid, α-tocopherol, flavonoids, polyamines) contents were significantly enhanced, accompanied by up-regulating the related enzyme and non-enzymatic antioxidant synthesis gene ( KcCu/Zn-SOD , KcAPX6 , KcHCT, KcHPT1 , Kcγ-TMT, KcF3H , KcSAMS and KcSMS ) expression with increasing concentrations of NaCl. These responses are beneficial for removing excess ROS to maintain a stable level of H 2 O 2 and O 2 - without lipid peroxidation in the K. caspia response to high salt. Meanwhile, up-regulating expression of KcSOS1/2/3, KcNHX1, and KcAVP was linked to Na + compartmentalization into vacuoles or excretion through salt glands in K. caspia . Notably, salt can improve the function of PSII that facilitate net photosynthetic rates, which is helpful to growing normally in high saline. Overall, the findings suggested that ROS scavenging systems and Na + /K + transport synergistically contributed to redox equilibrium, ion homeostasis, and the enhancement of PSII function, thereby conferring high salt tolerance., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Li, Mur, Wang, Hou, Zhao, Chen, Wu and Guo.)

Aim: The aim of this study was to understand the effects of revegetation on the diversity of bacteria and fungi in soil by sowing a single species and exploring the underlying mechanism., Location: Beijing, China., Taxon: Plants and Microbes., Methods: In a short-term ecological restoration experiment, one natural recovery treatment and three seed sowing treatments were chosen to assess their effects on the alteration of fungal and bacterial diversity. Plant species richness, abundance, and height were investigated. The diversity of fungi and bacteria was analyzed by high-throughput sequencing technologies. Linear mixed-effects model analysis was used to examine the effects of different restoration methods on biodiversity and ecosystem functions. Pearson's correlation analysis, analysis of covariance, and structural equation modeling (SEM) were used to examine the relationship between biodiversity and environmental factors., Results: Species richness and the Shannon-Wiener Index ( H ') of plants in the sown treatments were lower than in the natural recovery treatment, especially with sowing of Medicago sativa L. Similarly, the sum of the observed species and H ' of fungi and bacteria significantly decreased in the sown treatments. Moreover, plant density, community coverage, and soil moisture increased markedly, while soil bulk density decreased in the sown treatments. Importantly, SEM showed that sown treatments reduced the diversity of plants through increasing plant density, while it decreased the diversity of fungi and bacteria through decreasing the plant diversity and increasing soil moisture., Main Conclusions: Our findings confirm that ecological restoration by sowing could improve soil conditions, but may be unfavorable to the amelioration of soil microbial diversity in the short-term. Restoration practitioners should consider long-term studies on the dynamics of biodiversity in the above- and belowground after revegetation by native species to achieve goals related to biodiversity conservation., Competing Interests: None declared., (© 2019 The Authors. Ecology and Evolution published by John Wiley & Sons Ltd.)

Plant lignin is one of the major wall components that greatly contribute to biomass recalcitrance for biofuel production. In this study, total 79 representative Miscanthus germplasms were determined with wide biomass digestibility and diverse monolignol composition. Integrative analyses indicated that three major monolignols (S, G, H) and S/G ratio could account for lignin negative influence on biomass digestibility upon NaOH and H2SO4 pretreatments. Notably, the biomass enzymatic digestions were predominately affected by the non-KOH-extractable lignin and interlinked-phenolics, other than the KOH-extractable ones that cover 80% of total lignin. Furthermore, a positive correlation was found between the monolignols and phenolics at p<0.05 level in the non-KOH-extractable only, suggesting their tight association to form the minor wall-networks against cellulases accessibility. The results indicated that the non-KOH-extractable lignin-complex should be the target either for cost-effective biomass pretreatments or for relatively simply genetic modification of plant cell walls in Miscanthus.